US Army Medical Research and Development Command; Belgian
Walloon Region.

Background

There is currently no effective, practical vaccine against malaria. Protective
immunity has been achieved by irradiating infected Anopheles mosquitos
and then allowing them to bite subjects repeatedly. Although this is not
a feasible approach, it demonstrates the feasibility of producing protective
antibodies in the pre-erythrocytic stage of the disease, when sporozoites
are introduced into the body and migrate to the liver.

The circumsporozoite protein on the sporozoite is considered to be the
principal antigen responsible for these antibodies. In previous trials,
however, circumsporozoite vaccines have not been found to be sufficiently
immunogenic. In this trial, the authors report on a recombinant circumsporozoite
vaccine (consisting of the circumsporozoite protein fused to hepatitis
B surface antigen), in three different formulations with three different
adjuvants.

Methods

Subjects

Healthy volunteers, without prior exposure to malaria and without significant
medical problems.

Vaccine

The RTS,S vaccine, a recombinant polypeptide produced by yeast cells
and consisting of two components:

RTS is a polypeptide chain consisting of part of the circumsporozoite
protein fused to HBsAg.

S is the HBsAg polypeptide alone.

This vaccine was prepared in three different formulations, with different
adjuvants. Vaccine 1 consisted of RTS,S with alum and monophosphoryl lipid
A. Vaccine 2 was RTS,S in an oil-water emulsion. Vaccine 3 was RTS,S in
an oil-water emulsion plus the immune stimulants QS21 and monophosphoryl
lipid A.

The vaccines were administered i.m. at 0, 4 and 28 weeks. Thus, there
were three vaccines, each vaccine to be given in three doses.

Follow-up and measurements

Patients were monitored clinically at days 1, 2, 7 and 14 after each
vaccination.

Blood samples were obtained for the evaluation of humoral immunity on
the day of each vaccination, and at days 1 and 14 thereafter. Humoral immunity
was evaluted by determining antibodies to circumsporozoite protein (total
IgG levels and IgG subclass levels) and by an indirect fluorescence antibody
assay to air dried sporozoite. Antibodies to HBsAg were also determined.

Blood was obtained for the evaluation of cellular immunity (proliferative
and cytolytic assays) before the first vaccination and after the third
one.

Challenge with malaria

22 subjects who had received all three vaccine doses and 6 unimmunized
controls were challenged with falciparum malaria. This involved five bites
from laboratory-bred Anopheles mosquitos that had been infected with sporozoites.
Follow-up included frequent blood-smear examinations and treatment as needed
if infection developed.

Results

Subjects

46 subjects, men and women (mean age 31, range 18-45), were randomized
to one of the three vaccines in roughly equal numbers. The first dose was
well tolerated (mild local discomfort).

41 subjects went on to receive the second dose. Reactions to
vaccines 2 and 3 were more severe after the second dose (mainly constitutional
symptoms).

27 subjects received the third vaccine dose. The reasons for
not completing all three doses were mainly scheduling problems or noncompliance.
Because of the severity of the reaction after the second dose, two subjects
(one for vaccine 2 and one for vaccine 3) were not given the third dose
and the remainder received a reduced dose.

Of the 27 subjects who received all three doses, 22 agreed to
undergo sporozoite challenge (8 given vaccine 1 and 7 given vaccines 2
and 3).

Immunogenicity

Antibodies to circumsporozoite developed in all three vaccine groups,
but were higher in subjects who received vaccines 2 and 3. Similarly, subjects
who had been HBsAb negative before vaccination seroconverted, but antibody
response was higher in groups 2 and 3.

Vaccine efficacy

Sporozoite challenge in 6 unimmunized controls produced parasitemia
in all 6 after 11-13 days. 7 out of the 8 subjects given vaccine 1 became
parasitemic (12.6 days after exposure). 5 of the 7 subjects given vaccine
2 became parasitemic (after 15.2 days). Only 1 out of the 7 subjects given
vaccine 3 developed malaria, for a relative risk of infection of 0.14 (95%
CI 0.02 to 0.88).

Antibody titers were higher in subjects who were protected. Measurements
of cellular immunity did not clearly predict protection, however.

Authors' discussion

In the past, the only successful immunization against malaria had been
achieved using irradiated Anopheles mosquitos. This approach, although
impractical, showed that protective immunity to the pre-erythrocytic phase
of the disease (sporozoites) could be achieved and was the basis for the
development of the recombinant vaccine described here. The authors note
that although strong antibody response seems to be important, it is not
the only factor, since both vaccines 2 and 3 produced such responses but
only vaccine 3 was effective. They speculate that some effect of the adjuvant
in vaccine 3 on cellular immunity, perhaps on CD4+ T cells, may be responsible.

Although these results are very encouraging, they are preliminary, and
many more studies of safety and efficacy will be needed, particularly in
the field, in the setting of repeated exposure to multiple strains of plasmodium.

Editorial

In an editorial,
Nussenzweig and Zavala give an excellent summary of the work that has been
done so far in developing a malaria vaccine, of the problems that have
been overcome and the problems that remain to be dealt with.

Comment

Although the results of this study are very preliminary and may not be
borne out in practice, they are encouraging and fascinating. Malaria takes
a tremendous toll in lives and morbidity worldwide, and an effective vaccine
would be a major advance.

From a purely intellectual point of view, the scientific process leading
up to the vaccine described here and detailed in the editorial makes for
very interesting reading. For anyone without expertise in molecular biology
or immunology, reading the "methods" section of this article is a sobering
experience.